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Plasmid Files

pIEx™-2

Vector for high-level expression in insect cells of proteins with a cleavable N-terminal GST-6xHis-S-Tag cassette plus a C-terminal HSV tag.

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pIEx-2 Sequence and MappIEx-2.dna
Map and Sequence File   
Sequence Author:  Novagen (EMD Millipore)
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 DrdI (4095) PspFI (3897) BseYI (3893) AhdI (3309) BsaI (3243) BpmI (3240) NmeAIII (3162) AatII (2390) ZraI (2388) XbaI (2382) PflFI - Tth111I (2327) BspDI * - ClaI * (207) EcoRV (239) PsiI (409) NheI (606) BmtI (610) SphI (614) BsmI (775) BsiWI (784) MluI (872) BtgI - NcoI (1082) EcoNI (1097) EcoO109I (1119) MscI (1294) BsgI (1373) SwaI (1514) SpeI (1753) 6xHis XcmI (1786) PflMI (1830) PshAI (1904) BseRI (1908) BamHI (1926) MfeI (1932) Eco53kI (1941) BanII - SacI (1943) BsrGI (1946) AscI (1954) PstI - SbfI (1964) SalI (1966) Acc65I (1972) AgeI (1975) KpnI (1976) HindIII (1984) NotI (1991) BstZ17I (2006) PmlI (2011) BbsI (2045) PaeR7I - XhoI (2053) DraIII (2064) Bsu36I (2074) PacI (2103) pIEx™-2 4563 bp
DrdI  (4095)
1 site
G A C N N N N N N G T C C T G N N N N N N C A G

Sticky ends from different DrdI sites may not be compatible.
PspFI  (3897)
1 site
C C C A G C G G G T C G
BseYI  (3893)
1 site
C C C A G C G G G T C G

After cleavage, BseYI can remain bound to DNA and alter its
electrophoretic mobility.
AhdI  (3309)
1 site
G A C N N N N N G T C C T G N N N N N C A G

The 1-base overhangs produced by AhdI may be hard to ligate.
Sticky ends from different AhdI sites may not be compatible.
BsaI  (3243)
1 site
G G T C T C N C C A G A G N ( N ) 4

Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
BpmI  (3240)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI
recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its
electrophoretic mobility.
BpmI quickly loses activity at 37°C.
NmeAIII  (3162)
1 site
G C C G A G ( N ) 18-19 N N C G G C T C ( N ) 18-19

Efficient cleavage requires at least two copies of the NmeAIII
recognition sequence.
Sticky ends from different NmeAIII sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
AatII  (2390)
1 site
G A C G T C C T G C A G
ZraI  (2388)
1 site
G A C G T C C T G C A G
XbaI  (2382)
1 site
T C T A G A A G A T C T
PflFI  (2327)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (2327)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
BspDI  (207)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (207)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
EcoRV  (239)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to
delete a base after cleavage.
PsiI  (409)
1 site
T T A T A A A A T A T T
NheI  (606)
1 site
G C T A G C C G A T C G
BmtI  (610)
1 site
G C T A G C C G A T C G
SphI  (614)
1 site
G C A T G C C G T A C G
BsmI  (775)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
BsiWI  (784)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
MluI  (872)
1 site
A C G C G T T G C G C A
BtgI  (1082)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
NcoI  (1082)
1 site
C C A T G G G G T A C C
EcoNI  (1097)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
EcoO109I  (1119)
1 site
R G G N C C Y Y C C N G G R

Sticky ends from different EcoO109I sites may not be compatible.
MscI  (1294)
1 site
T G G C C A A C C G G T
BsgI  (1373)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14

Efficient cleavage requires at least two copies of the BsgI
recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
SwaI  (1514)
1 site
A T T T A A A T T A A A T T T A

SwaI is typically used at 25°C, but is 50% active at 37°C.
SpeI  (1753)
1 site
A C T A G T T G A T C A
XcmI  (1786)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
PflMI  (1830)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
PshAI  (1904)
1 site
G A C N N N N G T C C T G N N N N C A G

PshAI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
BseRI  (1908)
1 site
G A G G A G ( N ) 8 N N C T C C T C ( N ) 8

Sticky ends from different BseRI sites may not be compatible.
BseRI quickly loses activity at 37°C.
Prolonged incubation with BseRI may lead to degradation of the
DNA.
BamHI  (1926)
1 site
G G A T C C C C T A G G

After cleavage, BamHI-HF™ (but not the original BamHI) can
remain bound to DNA and alter its electrophoretic mobility.
MfeI  (1932)
1 site
C A A T T G G T T A A C
Eco53kI  (1941)
1 site
G A G C T C C T C G A G
BanII  (1943)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
SacI  (1943)
1 site
G A G C T C C T C G A G
BsrGI  (1946)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
AscI  (1954)
1 site
G G C G C G C C C C G C G C G G
PstI  (1964)
1 site
C T G C A G G A C G T C
SbfI  (1964)
1 site
C C T G C A G G G G A C G T C C
SalI  (1966)
1 site
G T C G A C C A G C T G
Acc65I  (1972)
1 site
G G T A C C C C A T G G
AgeI  (1975)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
KpnI  (1976)
1 site
G G T A C C C C A T G G
HindIII  (1984)
1 site
A A G C T T T T C G A A
NotI  (1991)
1 site
G C G G C C G C C G C C G G C G
BstZ17I  (2006)
1 site
G T A T A C C A T A T G
PmlI  (2011)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
BbsI  (2045)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
PaeR7I  (2053)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (2053)
1 site
C T C G A G G A G C T C
DraIII  (2064)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
Bsu36I  (2074)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
PacI  (2103)
1 site
T T A A T T A A A A T T A A T T
AmpR
2522 .. 3382  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   2522 .. 2590  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2522 .. 3382  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   2591 .. 3382  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2522 .. 3382  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
GST
1084 .. 1740  =  657 bp
219 amino acids  =  25.6 kDa
Product: glutathione S-transferase from
Schistosoma japonicum
GST
1084 .. 1740  =  657 bp
219 amino acids  =  25.6 kDa
Product: glutathione S-transferase from
Schistosoma japonicum
IE1 promoter
487 .. 1078  =  592 bp
promoter of the ie1 gene from the baculovirus
Autographa californica
IE1 promoter
487 .. 1078  =  592 bp
promoter of the ie1 gene from the baculovirus
Autographa californica
ori
3553 .. 4141  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3553 .. 4141  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
hr5 enhancer
1 .. 483  =  483 bp
baculovirus early transcription enhancer
hr5 enhancer
1 .. 483  =  483 bp
baculovirus early transcription enhancer
IE1 terminator
2080 .. 2387  =  308 bp
terminator of the ie1 gene from the baculovirus
Autographa californica
IE1 terminator
2080 .. 2387  =  308 bp
terminator of the ie1 gene from the baculovirus
Autographa californica
MCS
1900 .. 2014  =  115 bp
multiple cloning site
MCS
1900 .. 2014  =  115 bp
multiple cloning site
AmpR promoter
2417 .. 2521  =  105 bp
AmpR promoter
2417 .. 2521  =  105 bp
S-Tag
1795 .. 1839  =  45 bp
15 amino acids  =  1.7 kDa
Product: affinity and epitope tag derived from
pancreatic ribonuclease A
S-Tag
1795 .. 1839  =  45 bp
15 amino acids  =  1.7 kDa
Product: affinity and epitope tag derived from
pancreatic ribonuclease A
HSV tag
2020 .. 2052  =  33 bp
11 amino acids  =  1.2 kDa
Product: HSV (herpes simplex virus) epitope tag
HSV tag
2020 .. 2052  =  33 bp
11 amino acids  =  1.2 kDa
Product: HSV (herpes simplex virus) epitope tag
6xHis
1768 .. 1785  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
1768 .. 1785  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
thrombin site
1855 .. 1872  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
thrombin site
1855 .. 1872  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
enterokinase site
1891 .. 1905  =  15 bp
5 amino acids  =  606.5 Da
Product: enterokinase recognition and cleavage site
enterokinase site
1891 .. 1905  =  15 bp
5 amino acids  =  606.5 Da
Product: enterokinase recognition and cleavage site
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